DPPH Antioksidan Analizinin Yeniden Değerlendirilmesi

Antioksidanların uygulamaları, oksidatif stresin zararlı etkilerini en aza indirmedeki çeşitli rolleri nedeniyle artmaktadır. DPPH· (1,1-Difenil-2-pikrilhidrazil radikali; C18H12N5O6) antioksidan analizi, farklı bileşiklerin antioksidan aktivitesini değerlendirmek için yaygın olarak kullanılan hızlı, basit ve ucuz bir yöntemdir. Ayrıntılı literatür taraması, çeşitli araştırma gruplarının farklı DPPH· derişimlerinde (22,5–400µM), inkübasyon süresinde (5 dk-120 dk), reaksiyon çözücüsünde (etanol, metanol) ve reaksiyon karışım pH'sında (pH= 3 ve 5.5 tampon ortamı) olan oldukça farklı protokoller kullandıklarını göstermektedir. Farklı laboratuvarların sonuçlarının karşılaştırılması ve sonuçların doğrulanması için standart bir analizin gerekliliği çok önemlidir. Bu çalışmada DPPH· antioksidan analizini etkileyen DPPH· derişimi, standart madde derişimi, reaksiyon/inkübasyon süresi gibi önemli faktörler spektrofotometrik yöntem ile incelenerek standart bir DPPH· antioksidan analiz yöntemi oluşturulmuştur. Bu yöntem kapsamında ölçümler, reaksiyon ortamındaki DPPH· derişimi 25-75 µM aralığında, çalkalayıcılı inkübatörde 30oC’de, 30dk’da ve karanlık ortamda gerçekleştirilmelidir.

Anahtar Kelimeler:

DPPH, Antioksidan, Antioksidan aktivite, Yöntem

Re-Evaluating of The DPPH• Antioxidant Assay

Applications of antioxidants are increasing due to their various roles in minimising harmful effects of oxidative stress. DPPH· (1,1-Diphenyl-2-picrylhydrazyl radical) antioxidant assay is a fast, simple, inexpensive method commonly used to evaluate the antioxidant activity of different compounds. A detailed literature survey shows that various research groups have used widely different protocols differed in the concentration of DPPH· (22,5–400µM), incubation time (5 min–120 min), reaction solvent (ethanol, methanol) and pH of the reaction mixture (pH=3 and 5.5 buffer) . The requirement of a standard assay is very important in order to compare the results of different laboratories and validation of the conclusions. In this study, a standard DPPH• antioxidant analysis procedure was performed by analyzing the important factors such as DPPH• concentration, standard substance concentration, reaction / incubation time by spectrophotometric method. Under this procedure, the measurements should be carried out at DPPH concentration in the reaction medium in the range of 25-75 µM, 30 ° C in a shaker incubator, 30 minutes and dark medium.

Keywords:

DPPH• radical, Antioxidant, Antioxidant activity, Mthod,

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